Circ-TFF1 Promotes Breast Cancer Progression Through the miR-129-2-3p/IRAK1 Axis.

Breast cancer (BC) is a common malignant tumor, and circular RNA-trefoil factor 1 (circ-TFF1; hsa_circ_0061825) has been found to be highly expressed in BC tissues and cells and is associated with the poor prognosis of BC patients. However, the interaction between circ-TFF1 and microRNA in BC has not been studied. Quantitative real-time PCR was used to detect the expression of circ-TFF1, miR-129-2-3p, and interleukin (IL)-1 receptor-associated kinase 1 (IRAK1). Through the detection of cell proliferation, migration, invasion, tube formation, and apoptosis, cell function was assessed. The expression levels of angiogenesis-related proteins were detected by western blot. The interaction between miR-129-2-3p and circ-TFF1 or IRAK1 was verified by dual-luciferase reporter assay and RNA immunoprecipitation assay. Xenotransplantation experiments were used to confirm the function of circ-TFF1 in vivo. Circ-TFF1 and IRAK1 were significantly high expressed in BC tissues and cells. Silencing of circ-TFF1 reduced the proliferation, migration, invasion and tube formation, while increased the apoptosis of MDA-MB-361 and SK-Br-3 cells. MiR-129-2-3p was a target of circ-TFF1. Silencing of circ-TFF1 inhibited the malignant behavior of BC cells by releasing miR-129-2-3p. In addition, IRAK1 was a target of miR-129-2-3p. Overexpression of IRAK1 partially restored the inhibitory effect of miR-129-2-3p on cell progression. Animal experiments confirmed the anti-tumor effect of circ-TFF1 knockdown in vivo. Circ-TFF1 regulated the expression of IRAK1 by sponging miR-129-2-3p, thereby, promoting the development of BC. These data provided a novel targeted therapy for BC.

miR-129-2-3p directly targets SYK gene and associates with the risk of ischaemic stroke in a Chinese population.

Spleen tyrosine kinase (SYK) gene has been identified as novel susceptibility locus for ischaemic stroke (IS) previously. However, regulation of SYK gene remains unknown in IS. In this study, we aimed to identify miRNAs that might be involved in the development of IS by targeting SYK gene. miRNAs were firstly screened by bioinformatics predicting tool. The expression levels of SYK gene were detected by qRT-PCR and western blotting, respectively, after miRNA transfection. Luciferase reporter assay was applied to investigate the direct binding between miRNAs and target gene. miRNA levels were detected by miRNA TaqMan assays in the blood cells of 270 IS patients and 270 control volunteers. Results suggest that SYK gene might be a direct target of miR-129-2-3p. The blood level of miR-129-2-3p was significantly lower in IS patients (P < 0.05), and negatively associated with the risk of IS (adjusted OR: 0.88; 95% CI: 0.80-0.98; P = 0.021) by multivariable logistic regression analysis. The blood levels of SYK gene were significantly higher in IS patients, and miR-129-2-3p expression was negatively correlated with mean platelet volume. In summary, our study suggests that miR-129-2-3p might be involved in the pathogenesis of IS through interrupting SYK expression and the platelet function, and further investigation is needed to explore the underlying mechanism.

miR-129-2-3p binds SEMA4C to regulate HCC development and inhibit the EMT.

BACKGROUND: Among primary liver cancers, HCC is the most prevalent. Small noncoding RNAs called miRNAs control the expression of downstream target genes to take part in a variety of physiological and pathological processes, including those related to cancer. METHODS: miR-129-2-3p and SEMA4C expression levels were assessed using RT-qPCR. The CCK-8, invasion, and wound healing assays were used to confirm the capacity of HCC cells for proliferation, invasion and migration respectively. Serum SEMA4C levels were detected via ELISA. The RIP and dual-luciferase reporter assays were used to confirm the existence of intergenic binding sites. Cell apoptosis assay and cell cycle assay were performed to detect the apoptosis rate and cycle distribution of cells, and WB was performed to detect the protein expression of SEMA4C, RhoA, ROCK1, E-cadherin, N-cadherin, and vimentin. Furthermore, cancer-inhibiting role of miR-129-2-3p were further confirmed by animal tests. RESULTS: miR-129-2-3p expression was reduced in HCC tissues and cells. Overexpression of miR-129-2-3p decreased the proliferation, invasion, migration, and EMT in HCC cells, whereas inhibition of miR-129-2-3p had the opposite effects. Our research also showed that SEMA4C was increased in HCC tissues, serum and cells, and that SEMA4C knockdown prevented HCC cell invasion, migration, proliferation, and EMT. Overexpression of SEMA4C reversed the inhibitory effect of miR-129-2-3p on HCC. CONCLUSIONS: Overall, we discovered that through binding to SEMA4C, miR-129-2-3p regulates HCC cell proliferation, invasion, migration, and EMT.

miR-129-2-3p inhibits colon cancer cell proliferation by down-regulating the expression of BZW1.

BACKGROUND AND STUDY AIMS: MicroRNA (miRNA) is involved in diverse biological and physiological processes of tumors. Dysregulation of miRNA will induce a series of human diseases. miR-129-2-3p has vital effects in the pathogenesis of various tumors. However, the regulatory function of miR-129-2-3p in colon cancer remains to be clarified. This study investigated the role of miR-129-2-3p targeting BZW1 in proliferation, apoptosis, migration, and invasion of colon cancer. PATIENTS AND METHODS: Here, RT-qPCR was applied to measure the miR-129-2-3p levels in colon cancer tissues. The predicted targets of miR-129-2-3p were identified by bioinformatics and verified using luciferase reporter assay. The effects of miR-129-2-3p on colon cancer were detected by CCK-8, colony formation, transwell chamber test, wound healing, and flow cytometry assays. Finally, the influence of miR-129-2-3p on tumor growth was studied. Nude mice were xenografted with transfected Lovo cells by subcutaneous injection of 5 × 105 cells in 100 µl. HE staining and TUNEL were used to assess metastasis ability. RESULTS: miR-129-2-3p level in colon cancer tissue was significantly reduced. Furthermore, it was verified that BZW1 was a target of miR-129-2-3p, and its expression in colon cancer cells was inhibited by miR-129-2-3p. Additionally, miR-129-2-3p inhibited colon cancer cell proliferation, colony formation, mobility ability and tumor growth, and promoted cell apoptosis by targeting BZW1. miR-129-2-3p overexpression in tumor xenografts in vivo decreased BZW1 expression, and suppressed tumor growth. CONCLUSION: Collectively, these findings indicated that miR-129-2-3p exerts a suppressive role in colon cancer cells by directly targeting BZW1, and may have significant therapeutic implications for patients with colon cancer.

miR-129-2-3p mediates LPS-induced macrophage polarization and ferroptosis by targeting the SMAD3-GPX4 axis.

Macrophages has become a promising target of sepsis treatment because macrophages dysfunction contributes to the progress of sepsis. The targeted therapy of sepsis based on macrophages ferroptosis is drawing more and more attention, but the molecular mechanism involved is poorly understood. In this study, Mus musculus-derived macrophages were used for in-vitro experiments. We found that LPS could induce ferroptosis in macrophages via the detection of apoptosis, GSH, lipid peroxide and GPX4 levels. Meanwhile, miR-129-2-3p was up-regulated in macrophages exposure to LPS. Next, we confirmed that miR-129-2-3p promoted the LPS-induced polarization of M1 phenotype in macrophages via the detection of Arg-1 and iNOS levels; miR-129-2-3p promoted the LPS-induced ferroptosis in macrophages. Further, luciferase assay showed that SMAD3 was identified as a target gene of miR-129-2-3p and its expression was negatively regulated by miR-129-2-3p and LPS. SMAD3 could inhibit the LPS-induced polarization of M1 phenotype and ferroptosis in macrophages by targeting GPX4. Collectively, we demonstrated the target gene and molecular mechanism of miR-129-2-3p mediating LPS-induced polarization and ferroptosis in macrophages by targeting the SMAD3-GPX4 axis, which would provide a novel strategy for sepsis targeted therapy based on macrophages polarization and ferroptosis.

Exosomal-miR-129-2-3p derived from Fusobacterium nucleatum-infected intestinal epithelial cells promotes experimental colitis through regulating TIMELESS-mediated cellular senescence pathway.

Fusobacterium nucleatum (Fn) infection is known to exacerbate ulcerative colitis (UC). However, the link between Fn-infected intestinal epithelial cell (IEC)-derived exosomes (Fn-Exo) and UC progression has not been investigated. Differentially expressed miRNAs in Fn-Exo and non-infected IECs-derived exosomes (Con-Exo) were identified by miRNA sequencing. Then, the biological role and mechanism of Fn-Exo in UC development were determined in vitro and in vivo. We found that exosomes delivered miR-129-2-3p from Fn-infected IECs into non-infected IECs, exacerbating epithelial barrier dysfunction and experimental colitis. Mechanically, Fn-Exo induces DNA damage via the miR-129-2-3p/TIMELESS axis and subsequently activates the ATM/ATR/p53 pathway, ultimately promoting cellular senescence and colonic inflammation. In conclusion, Exo-miR-129-2-3p/TIMELESS/ATM/ATR/p53 pathway aggravates cellular senescence, barrier damage, and experimental colitis. The current study revealed a previously unknown regulatory pathway in the progression of Fn-infectious UC. Furthermore, Exosomal-miR-129-2-3p in serum and TIMELESS may function as novel potential diagnostic biomarkers for UC and Fn-high-UC.

MiR-129-2-3p Inhibits Esophageal Carcinoma Cell Proliferation, Migration, and Invasion via Targeting DNMT3B.

PURPOSE: The study aims to explore the regulatory mechanism of miR-129-2-3p underlying esophageal carcinoma (EC) cell progression and generate new ideas for targeted treatment of EC. METHODS: Mature miRNA expression data and total RNA sequencing data of EC in the TCGAESCA dataset were utilized to explore differentially expressed miRNAs (DEmiRNAs). StarBase database was then utilized to predict targets of miRNA. MiR-129-2-3p and DNMT3B expression in EC cell lines was assayed through qRT-PCR and Western blot. CCK-8, scratch healing, and transwell assays were conducted to assess the impact of miR-129-2-3p on EC cell phenotypes. In addition, a dual-luciferase assay was completed to identify the binding relationship between DNMT3B and miR-129-2-3p. RESULTS: MiR-129-2-3p was noticeably less expressed in EC cell lines, while DNMT3B was highly expressed. MiR-129-2-3p could bind to DNMT3B. Furthermore, in vitro functional experiments uncovered that overexpressed miR-129-2-3p repressed EC cell progression while further overexpressing DNMT3B would restore the above inhibitory effect. CONCLUSION: MiR-129-2-3p is a cancer repressor in EC cells, and it could target DNMT3B, thus hampering the progression of EC cells.

Rolling circle amplification (RCA) -based biosensor system for the fluorescent detection of miR-129-2-3p miRNA.

Herein, a versatile fluorescent bioanalysis platform for sensitive and specific screening of target miRNA (miR-129-2-3p) was innovatively designed by applying target-induced rolling circle amplification (RCA) for efficient signal amplification. Specifically, miR-129-2-3p was used as a ligation template to facilitate its ligation with padlock probes, followed by an RCA reaction in the presence of phi29 DNA polymerase. The dsDNA fragments and products were stained by SYBR Green I and then detected by fluorescence spectrophotometry. As a result, miR-129-2-3p concentrations as low as 50 nM could be detected. Furthermore, the expression of miR-129-2-3p in breast cancer patients was about twice that in healthy people. Therefore, the results indicated that the RCA-based biosensor system could be a valuable platform for miRNA detection in clinical diagnosis and biomedical study.

Circ_0081143 Contributes to Gastric Cancer Malignant Development and Doxorubicin Resistance by Elevating the Expression of YES1 by Targeting mziR-129-2-3p.

Background/Aims: The increased mortality of gastric cancer (GC) is mainly attributed to the development of chemoresistance. Circular RNAs, as the novel type of biomarkers in GC, have attracted wide attention. The purpose of this study was to investigate the functional role of circ_0081143 in GC with doxorubicin (DR) resistance and its potential action mechanism. Methods: The expression of circ_0081143, miR-129-2-3p and YES proto-oncogene 1 (YES1) in GC tissues and cells was measured by quantitative real-time polymerase chain reaction. The half maximal inhibitory concentration value was calculated based on the MTT cell viability assay. Cell proliferation and apoptosis were monitored by MTT and flow cytometry assays. Transwell assays were employed to check cell migration and invasion. The protein levels of YES1 and apoptosis-related proteins were detected by western blotting. The interaction between miR-129-2-3p and circ_0081143 or YES1 was verified by dual-luciferase reporter and pull-down assays. A tumorigenicity assay was conducted to verify the role of circ_0081143 in vivo. Results: Circ_0081143 was highly expressed in DR-resistant GC tumor tissues and cells. Depletion of circ_0081143 reduced DR resistance and inhibited DR-resistant GC cell proliferation, migration and invasion. Circ_0081143 targeted miR-129-2-3p and inhibited the role of miR-129-2-3p. In addition, YES1 was a target of miR-129-2-3p, and its function was suppressed by miR-129-2-3p. Importantly, circ_0081143 positively modulated the expression of YES1 through mediating miR-129-2-3p. Circ_0081143 knockdown weakened the DR-resistant GC tumor growth in vivo. Conclusions: Circ_0081143 knockdown weakened DR resistance and blocked the development of DR-resistant GC by regulating the miR-129-2-3p/YES1 axis. Our data suggest that circ_0081143 is a promising target for the treatment of GC with DR resistance.

Long Noncoding RNA LINC01006 Facilitates Cell Proliferation, Migration, and Epithelial-Mesenchymal Transition in Lung Adenocarcinoma via Targeting the MicroRNA 129-2-3p/CTNNB1 Axis and Activating Wnt/ß-Catenin Signaling Pathway.

Lung adenocarcinoma (LUAD) is a common type of malignancy of lung cancers. Long intergenic noncoding RNAs (lincRNAs) have emerged as crucial regulators of various cancers, including LUAD. LINC01006 is a newly discovered long noncoding RNA (lncRNA) whose function in LUAD remains to be explored. This study is to explore the role of LINC01006 in LUAD. Quantitative real-time PCR (RT-qPCR) analysis and Western blotting were used to determine the expression levels and protein levels, respectively. Functional assays and animal experiments investigated the role of LINC01006 both in vivo and in vitro. Moreover, TOP/FOP assay was performed to detect the activation of the Wnt/ß-catenin signaling pathway. The interaction between LINC01006 and microRNA 29-2-3-p (miR-29-2-3-p)/catenin beta 1 (CTNNB1) was explored by RNA binding protein immunoprecipitation (RIP), RNA pulldown, luciferase reporter assays, and rescue experiments. According to the results, LINC01006 was highly expressed in LUAD tissues and cell lines. LINC01006 knockdown significantly suppressed cell proliferative, migratory, and epithelial-mesenchymal transition (EMT) capacities and tumor development. Moreover, LINC01006 enhanced CTNNB1 via sequestering miR-129-2-3p and activated the Wnt/ß-catenin pathway in LUAD. Overall, LINC01006 promotes LUAD development via activating the Wnt/ß-catenin pathway, implying that LINC01006 might be a promising biomarker for LUAD treatment.

Inhibition of microRNA-129-2-3p protects against refractory temporal lobe epilepsy by regulating GABRA1.

BACKGROUND: Accumulating evidence demonstrates that certain microRNAs play critical roles in epileptogenesis. Our previous studies found microRNA (miR)-129-2-3p was induced in patients with refractory temporal lobe epilepsy (TLE). In this study, we aimed to explore the role of miR-129-2-3p in TLE pathogenesis. METHOD: By bioinformatics, we predicted miR-129-2-3p may target the gene GABRA1 encoding the GABA type A receptor subunit alpha 1. Luciferase assay was used to investigate the regulation of miR-129-2-3p on GABRA1 3'UTR. The dynamic expression of miR-129-2-3p and GABRA1 mRNA and protein levels were measured in primary hippocampal neurons and a rat kainic acid (KA)-induced seizure model by quantitative reverse transcription-polymerase chain reaction (qPCR), Western blotting, and immunostaining. MiR-129-2-3p agomir and antagomir were utilized to explore their role in determining GABRA1 expression. The effects of targeting miR-129-2-3p and GABRA1 on epilepsy were assessed by electroencephalography (EEG) and immunostaining. RESULTS: Luciferase assay, qPCR, and Western blot results suggested GABRA1 as a direct target of miR-129-2-3p. MiR-129-2-3p level was significantly upregulated, whereas GABRA1 expression downregulated in KA-treated rat primary hippocampal neurons and KA-induced seizure model. In vivo knockdown of miR-129-2-3p by antagomir alleviated the seizure-like EEG findings in accordance with the upregulation of GABRA1. Furthermore, the seizure-suppressing effect of the antagomir was partly GABRA1 dependent. CONCLUSIONS: The results suggested GABRA1 as a target of miR-129-2-3p in rat primary hippocampal neurons and a rat kainic acid (KA) seizure model. Silencing of miR-129-2-3p exerted a seizure-suppressing effect in rats. MiR-129-2-3p/GABRA1 pathway may represent a potential target for the prevention and treatment of refractory epilepsy.

MicroRNA-129-2-3p directly targets Wip1 to suppress the proliferation and invasion of intrahepatic cholangiocarcinoma.

Accumulated studies showed that numerous microRNAs (miRNAs) were aberrantly expressed in human intrahepatic cholangiocarcinoma (ICC) and contributed to the tumorigenic processes. However, whether miR-129-2-3p is implicated in the ICC initiation and progression is still limited. Here, the results revealed that miR-129-2-3p expression was notably decreased in ICC tissues and cell lines, and that a low miR-129-2-3p expression was obviously associated with distant metastasis and clinical stage. Exogenous miR-129-2-3p expression evidently repressed the proliferative and invasive abilities of ICC cells. Mechanistic studies indicated that Wild-type p53-induced phosphatase 1 (Wip1) was a direct target gene for miR-129-2-3p in ICC cells. Furthermore, silencing Wip1 expression mimicked the suppressive effects of miR-129-2-3p upregulation on ICC cells. Interestingly, reintroduction of Wip1 expression partially abolished the miR-129-2-3p -reduced cell proliferation and invasion in ICC. Moreover, ectopic miR-129-2-3p expression hindered the ICC tumor growth in vivo. To the best of our knowledge, it is the first time to reveal that miR-129-2-3p plays a crucial role in tumor suppression in ICC pathogenesis through directly targeting Wip1. These results will aid in elucidating the roles of miR-129-2-3p in ICC, and suggest that this miRNA may provide a potential target for the treatment of ICC.

Mir-129-2-3p Has Tumor Suppressor Role in Ewing Sarcoma Cell Lines and Cancer Tissue Samples

Abstract Deregulation of miRNA expressions was identified as a novel feature of tumor biology in Ewing sarcoma (EWS). The aim was to evaluate the regulatory role of miR-129-2-3p in EWS cell lines and human EWS tissue samples. EWS cell lines TC-71, TC-106, and CHLA-99 were used in the study and real-time PCR was utilized to investigate the functional role of tumor suppressor mir-129-2-3p and miR-129-2-3p levels in the cells. Proliferation, migration, invasion and apoptosis assays were carried out within the scope of functional in vitro studies. Expression levels of CDK6 and SOX4, which are miR-129-2-3p target genes, were examined. Moreover, the change in expression levels of miR-129-2-3p in EWS tumor tissues was also examined. It was determined that miR-129-2-3p expression markedly diminished in all the studied cell lines. In addition, miR-129-3p was found to decrease in proliferation, migration, invasion and apoptosis assays in all EWS cell lines. CDK6 and SOX4 levels were also decreased in miR-129-2-3p transfected cell lines. It was found that miR-129-2-3p levels were significantly decreased in EWS tumor tissue samples compared to the corresponding adjacent normal tissue samples. In line with the results of our current study, where the possible function of miR‐129-2-3p in EWS cell lines was examined, for the first time in the literature miR-129-2-3p was shown to have low expression level in EWS lines and EWS tumor tissue samples, and to provide a tumor suppressor effect.